Fixing Off Target Effects of CRISPR/Cas9
CRISPR’s simplicity and accessibility has given itself the oxygen to spread like wildfire through the research community, but so have it’s off target effects. Recently, scientists have used it in clinical trials for slowing or stopping the production of lung cancer cells, but due to these concerns about its off target effects of CRISPR/Cas9, scientists are wondering how effective it will actually be in humans. This leads to another major question and concern, what the off target effects will be in human trials.
Potential/Questionable Mutant Conclusions
It was determined by Kellie A. Schaefer and collaborators at Stanford that the accuracy of CRISPR/Cas9 in regard to its off-target mutations, must be further improved, particularly when referring to clinical therapies. Kellie A. Schaefer, Ph.D., at Stanford University and colleagues at Howard Hughes and Massachusetts General Hospital conducted whole genome sequencing (WGS) on DNA isolated from two CRISPR-repaired mice (F03 and F05) and one uncorrected control. They said the sequencing found was of an exceptionally high value of single nucleotide variations (SNVs) in comparison to the average assumption that CRISPR causes mutations primarily at regions homologous to the sgRNA. Benjamin P. Kleinstiver Ph.D. of the Molecular Pathology Unit and Center for Cancer Research at the Massachusetts General Hospital and other investigators hypothesized that reducing the contact between iCas9 and targeted DNA might help reduce the off-target effects, without compromising the on-target interactions. In the BioRxiv paper by Wilson et al. it was noted the Schaefer et al. study was technically inconclusive due to small sample size. Curiously, one of Wilson’s co-authors for BioRxiv, George Church, has a vested interest in the success of CRISPR technologies. Click to learn more about a CRISPR knockout.